SiiA and SiiB are novel type I secretion system subunits controlling SPI 4‐mediated adhesion of S almonella enterica

Summary The giant non‐fimbrial adhesin SiiE is essential to establish intimate contact between S almonella enterica and the apical surface of polarized epithelial cells. SiiE is secreted by a type I secretion system ( T 1 SS ) encoded by S almonella   P athogenicity I sland 4 ( SPI 4). We identified SiiA and SiiB as two regulatory proteins encoded by SPI 4. Mutant strains in siiA or siiB still secrete SiiE , but are highly reduced in adhesion to, and invasion of polarized cells. SiiA and SiiB are inner membrane proteins with one and three transmembrane ( TM ) helices respectively. TM 2 and TM 3 of SiiB are similar to members of the ExbB / TolQ family, while the TM of SiiA is similar to MotB and a conserved aspartate residue in this TM is essential for SPI 4‐encoded T 1 SS function. Co‐immunoprecipitation, bacterial two‐hybrid and FRET demonstrate homo‐ and heterotypic protein interactions for SiiA and SiiB . SiiB , but not SiiA also interacts with the SPI 4‐ T 1 SS ATP ase SiiF . The integrity of the W alker A box in SiiF was required for SiiB – SiiF interactionand SiiF dimer formation. Based on these data, we describe SiiA and SiiB as new, exclusively virulence‐associated members of the Mot / Exb / Tol family of membrane proteins. Both proteins are involved in a novel mechanism of controlling SPI 4‐ T 1 SS ‐dependent adhesion, most likely by formation of a proton‐conducting channel.